LIGHT-DRIVEN ION TRANSPORT IN BACTERIAL RHODOPSINS

超出提供的空间。细菌视紫红质是最简单的离子泵，也是普遍存在的七螺旋膜蛋白的结构原型，在过去的几年里，我们测定了这个光驱动质子传输系统中七个捕获的光中间体的X射线衍射结构。其余两个态的结构可以间接地从非发光突变体获得。视网膜和视蛋白的构象变化暗示了这种泵的原子水平机制，但如果没有所有九种中间体的结构，这种机制是不完整的。直接信息较少的状态是跨膜螺旋的大范围运动调节潜水分子的进出，影响质子供体和受体的PKS，形成质子传导链。这些状态不能被捕获在三维晶体中。我们将专注于通过定点定向自旋标记来测试和验证运输周期的这些关键状态的结构。大量位于战略位置的胱氨酸残基将揭示突变结构与N，N‘和O中间体的对应程度，如果不同，则开始定义这些状态的结构。同时，关于俘获状态的结晶学研究将继续改进现有的结构，并使用可见光和红外光谱来探索关于蛋白质胞外区域质子转移的仍然悬而未决的问题。预计这项工作的结果将是首次全面描述活性部位的局部变化与任何转运体或受体中蛋白质的整体变化的耦合。表演网站========================================Section End===========================================

EXCEED THE SPACE PROVIDED. Bacteriorhodopsin is the simplest of ion pumps, and the structural prototype of the ubiquitous heptahelical membrane proteins, in the last few years we determined the x-ray diffraction structures of seven trapped photointermediates of this light-driven proton transport system. The structures of the remaining two states are available indirectly from non-illuminated mutants. The conformational transformations of the retinal and opsin suggest an atomic level mechanism for this pump, but the mechanism is incomplete without the structures of all nine intermediates. The states with less direct information are those where large-scale motions of the transmembrane helices mediate the entry and exit of buried water molecules that affect the pKs of proton donors and acceptors and form proton conducting chains. These states cannot be trapped in 3-dimensional crystals. We will focus on testing and authenticating the structures of these crucial states of the transport cycle by site-directed spin-labeling. A large number of strategically located cystine residues will reveal how well the mutant structures correspond to the N, N' and O intermediates, and if different, begin to define the strucures of these states. At the same time, crystallographic studies with trapped states will continue to improve the structures already available, and to use visible and infrared spectroscopy to explore still outstanding questions concerning proton transfers in the extracellular region of the protein. The outcome of this work is expected to be the first full description of the coupling of local changes at the active site to global changes of the protein in any transporter or receptor. PERFORMANCE SITE ========================================Section End===========================================